Why are my E. coli cells not pellet-able?

I'm expressing a lipopolysaccharide biosynthetic enzyme in E. coli. This is a enzyme from C. burnetii chimerized with a short C-terminal sequence from its E. coli homolog. The recombinant protein expressed as cytosolic oluble protein in E. coli BL21(DE3). But interestingly, comparing to the wild-type C. burbetii enzyme, this protein seems to make the E. coli cells unable to pellet well.

The cell are being pelleted by a centrifuge. Stickiness / clumping... while a factor isn't the only one. The density of the cells and the number (thus the pellet size, after x time of centrifuging) also play a role.

You could try increasing the centrifuge speed. Or spin for longer.

May your PCR products be long, your protocols short and your boss on holiday

We had a mutant that over-produced an exopolysaccharide. We could not, for the life of us, get this strain to pellet; this phenotype was actually how we found what proved to be a very interesting EPS locus and regulatory mechanism in our bug.

That's very interesting! Can you give me the citation of your paper published on that?Thank you for your input!

Lydia

We had a mutant that over-produced an exopolysaccharide. We could not, for the life of us, get this strain to pellet; this phenotype was actually how we found what proved to be a very interesting EPS locus and regulatory mechanism in our bug.